Interleukin 1 (IL-1) is a polypeptide cytokine with multiple diverse effects on immunological and inflamatory processes. While many of the roles of IL-1 in inflammation and the immune response have been well characterized, the molecular basis of these responses remains unclear (reviewed by Dinarello, Blood, 77: 1627-1652). IL-1 is produced by a diversity of cell types and elicits a wide variety of physiological effects in hematopoetic and nonhematopoetic cells. Thus, IL-1 has biological effects on hematopoietic cells, the digestive tract, bone, cartilage and connective tissue, vascular cells, the skin, the endocrine system, the gonads, and on neural tissue. In addition, IL-1 is produced by malignant cells. (Pimentel, Handbook of Growth Factors: Volume III Hematopoietic Growth Factors and Cytokines, pp. 35-53, CRC Press, Boca Raton, Fla. 1994).
The IL-1 family of proteins comprises three members: IL-1 γ and IL-1β (capable of inducing IL-1 biological responses) and IL-1R (a pure receptor antagonist). These ligands bind to two distinct and separate receptors: the Type I and Type II IL-1 receptors (IL-1Rs). The 80-kD Type I IL-1R is found mainly on T cells and fibroblasts (Sims, J. E., et al., Science 241:585-589 (1988); Chizzonite, R., et al., Proc. Natl. Acad. Sci. USA 86:8029-8033 (1989); Sims, J. E., et al., Proc. Natl. Acad. Sci. USA 86:8946-8950 (1989)). The 68-kD Type II IL-1R is found predominantly on B cells and neutrophils (Chizzonite, R., et al., Proc. Natl. Acad Sci. USA 86:8029-8033 (1989); Sims, J. E., et al., Proc. Natl. Acad, Sci. USA 86:8946-8950 (1989); McMahan, C. J., et al., EMBO J. 10:2821-2832 (1991)). Both receptor types contain a large cytoplasmic region, a single transmembrane domain, and the extracellular Ig-like domains, a structural organization that classifies them as members of the Ig superfamily. The Type I IL-1R has a cytoplasmic tail of approximately 200 amino acids, while the Type II IL-1R cytoplasmic tail is only 29 amino acids. The agonists IL-1α and IL-1β bind to the extracellular domains of both receptors, although with different affinities (reviewed in Dower et al., Cellular and Molecular Mechanisms of Inflammation, pp. 137-172, Academic Press, Orlando Fla.).
The relative importance of the Type I and Type II IL-1Rs in IL-1 signaling has been recently clarified. A critical role for the Type I IL-1R in IL-1-induced activation of NF-kB, IL-6, and IL-8 secretion, and cell adhesion molecule expression has been demonstrated by several groups (Stylianou, E., et al., J. Biol. Chem 267:15836-15841 (1992); Colotta, F., et al., Science 261:472475 (1993); Sims, J. E., et al., Proc. Natl. Acad. Sci. USA 90:6155-6159 (1993)). In contrast, the Type II IL-1R appears to be dispensable for IL-1 signaling and may act as a decoy receptor (Stylianou, E., et al., J: Biol. Chem. 267:15836-15841 (1992); Colotta, F., et al., Science 261:472475 (1993); Sims, J. E., et al, Proc. Natl. Acad. Sci. USA 90:6155-6159 (1993)). While it appears clear that the Type I IL-1R is necessary for IL-1 signal transduction, it is uncertain if it is the only cell-surface molecule involved in IL-1 signaling.
It has been assumed that the functional Type I IL-1R is a single chain receptor (Curtis, B. M., et al., Proc. Natl. Acad. Sci. USA, 86:3045-3049 (1989)). However, affinity cross-linking of IL-1 to cells expressing natural IL-1 receptor has yielded complex patterns of cross-linked proteins (Dower, et al., Cellular and Molecular Mechanisms of Inflammation, pp. 137-172, Academic Press, Orlando Fla. (1990); Dinarello, et al., Immunol. Today, 10: 49-51 (1989)). These cross-linking studies detect molecular mass complexes consistent with both the Type I and Type II IL-1Rs cross-linked to IL-1. In addition, in some studies, higher molecular mass complexes (>200 kD) are apparent (Kupper, T. S., et al., J. Clin. Invest. 82:1787-1792 (1988); Dinarello, C. A., et al., Immunol. Today 10:49-51 (1989); Solari, R., Cytokine 2:21-28 (1990); Mancilla, J., et al., Lymph. Cytokine Res. 11:197-205 (1992)). Some reports have interpreted these higher molecular mass complexes to be dimers of receptor-ligand complexes. Others have concluded that these high molecular mass complexes may be indicative of a multi-subunit IL-1 receptor complex.
Only two IL-1R accessory proteins are have been identified. Studies initiated to identify components of a potential IL-1 receptor complex suggest that three is a cell-surface protein in close association with the IL-1R that may play a role in IL-1 receptor binding and signaling. A murine IL-1 receptor accessory protein (muIL-1R AcP) has been cloned and expressed (Greenfeder et al. J. Biol. Chem, 270: 13757-13765 (1995)). This protein was present in brain, lung, spleen, and thymus tissues. A search of the GenBank data base with the muIL-1R AcP cDNA sequence revealed significant homology (82%) to a cDNA isolated from human infant brain (accession no. T08277) (Adams, M. D., et al, Nature Genet. 4:373-380 (1993)). No other significant homologies were found in GenBank. The reported sequence for this partial cDNA is 396 bp long and represents one of 1600 cDNAs that were sequenced from a library made to contain only expressed sequence tags. The regions to overlap with the muIL-1R AcP sequence is nucleotides 893-1286 of the muIL-R AcP, which include the transmembrane domain. Although Adams et al. (Adams, M. D., et al., Nature Genet 4:373-380 (1993)) assigned no function to this partial cDNA, it is likely that it encodes a portion of a human homologue of mulIL-1R AcP. Using the mulIL-1R AcP cDNA has >95% homology to the partial sequence of Adams et al. and ˜90% homology to the mulL-1R AcP cDNA. This partial cDNA was isolated as an expressed gene in infant brain. This was consistent with Northern analysis results of Greenfeder et al. demonstrating that mulIL-1R AcP mRNA is constitutively expressed at high levels in mouse brain.
The discovery of IL-1R accessory molecule has a number of implications for IL-1 receptor biology. First, while muIL-1R AcM may not bind IL-1 directly, the accessory molecule forms a complex with the muType I IL-1R allowing IL-1β to bind with higher affinity than the muType I IL-1R alone (Greenfeder, et al. J. Biol. Chem, 270: 13757-13765 (1995)). Thus, the presence or absence of the accessory molecule in different cell lines determined whether the low or the higher affinity site was detected, suggesting that the low affinity site corresponds to the muType I IL-1R alone, while the higher affinity site represents a complex of the muType I IL-1R with the muIL-1R AcM. (Greenfeder, et al. J. Biol. Chem, 270:13757-13765 (1995)). In this respect, the IL-1R AcM would be analogous to affinity conversion and signal transduction subunits such as gp 130 in the IL-6 system (Hibi, M, et al., Cell 63:1149-1157 (1990)), the common β chain of the IL-3, granulocyte/macrophage colony-stimulating factor, and IL-5 receptors (Kitamura, T., et al., Cell 66:1165-1174 (1991)), and the γc subunit first identified as part of the IL-2 receptor (reviewed in Miami, et al., Annu. Rev. Immunol. 11: 245-267 (1993)).
Second, the possible existence of a multi-subunit IL-1 receptor complex contradicts a previous hypothesis that the Type I IL-1R is the entire functional receptor for IL-1 signaling (Dower, S. K. & Sims, J. E., Cellular and Molecular Mechanisms of Inflammation, Academic Press, Orlando, Fla. (1990), pp. 137-172; Curtis, B. M., et al., Proc. Natl. Acad. Sci USA, 86:3045-3049 (1989)). This hypothesis was based on the observation that CHO cells expressing recombinant murine Type I IL-1R were more sensitive than control CHO cells to low concentrations of IL-1, and that the increase in sensitivity was proportional to the number of murine Type I IL-1Rs (Curtis, B. M., et al., Proc. Natl. Acad Sci USA, 86:3045-3049 (1989)). An alternative explanation for these results is that the endogenous hamster IL-1R, was able to form a functional receptor complex with the mu Type I IL-1R, thus enhancing IL-1 signaling in the transfected cells (Greenfeder, et al. J. Biol. Chem, 270: 13757-13765 (1995)).
Third, the discovery of the accessory protein provides an intriguing explanation for the antagonist activity of IL-1ra despite its high affinity binding to the Type I IL-1R The inability of IL-1ra to interact with the muIL-1RAcP, the putative signal transducing subunit of the IL-1R complex, would result in the absence of a biological response.